Biphenyl Acetate, Preparation and Uses Thereof

ABSTRACT

A biphenyl acetate composition, which is a biphenyl acetic ammonia butantriol salt. The composition is prepared by reacting biphenyl acetic acid and ammonia butantriol in organic solvent, and a ratio of biphenyl acetic acid and ammonia butantriol is 1:1. The salt is capable of providing anti-inflammatory, analgesic, and antipyretic effect. The salt is unique and has stable chemical structure, can be prepared into an injection or a capsule, and has low toxicity and insignificant local and vascular irritating effect, thereby being prepared into a predetermined dosage in a controllable and precise manner.

CROSS REFERENCE OF RELATED APPLICATION

This is a continuation application that claims the benefit of priorityunder 35 U.S.C. §119 to a non-provisional application, application Ser.No. 12/309,326 with a filing date of Jan. 12, 2009, which is a nationalphase national application of an international patent application numberPCT/CN2007/002830 with a filing date of Sep. 27, 2007 based on a foreignapplication number 200610122917.4 with a filing date of Oct. 23, 2006 inChina. The contents of these specifications, including any interveningamendments thereto, are incorporated herein by reference.

BACKGROUND OF THE PRESENT INVENTION

1. Field of Invention

The present invention relates to preparation and use of biphenylacetate, and more particularly to a preparation and use of biphenylacetic ammonia butantriol salt.

2. Description of Related Arts

In 1993, biphenyl acetic acid is commonly used as a medicine forreducing inflammation and relieving pain in Japan and Italy. Thebiphenyl acetic acid is used to provide analgesia and anti-inflammatoryeffect related tometaboly arthritis, omarthritis, tenosynovitis, musclepain and cartilage tissue injury. In addition, it is also used forreducing traumatic swelling and pain. Because of its insolubility inwater, biphenyl acetate can only be used for traumatic medicine. Thisproperty restricts the application of this medicine dramatically.

In Japan, biphenyl ethyl acetate has been used for injecting emulsion(insoluble in water) for reducing clinical pain of a living object.However, the manufacture of this medication has recalled this type ofemulsion recently because it is discovered that the emulsion may cause ashock phenomenon to the living object. Further clinical studies foundthat allergy is the major cause of the shock phenomenon and the soyphosphatidoo acting as the coemulsifier of the injecting emulsion actsmay be one of the allergen. Therefore, the biphenyl ethyl acetate whichis manufactured in an emulsion form has the drawbacks of very lowstability, involving complicated preparation process and technique andbeing unsafe owing to the macromolecular structure of the coemulsifier.In other words, in view of the functional group of biphenyl acetic acid,there is no solution provided to make use of this functional group forpreparation of injection solution or the like which is stable instructure and safe for administration as injection to a living object.It is worth mentioning that the injecting emulsion containsmacromolecular protein which may not only impose toxic effect on liverbut also cause allergy to the living object, therefore imposing safetyissue when used in living object.

SUMMARY OF THE PRESENT INVENTION

An object of the present invention is to provide a new kind of biphenylacetate, which is biphenyl acetic ammonia butantriol salt. The biphenylacetic ammonia butantriol salt has the following chemical structure.

Its molecular structure is C₁₈H₂₃NO₅, and its molecular weight is333.14.

Another object of the present invention is to provide a manufacturingmethod of biphenyl acetic ammonia butantriol salt. This method is toreact biphenyl phenylacetic acid with ammonia butantriol in organicsolvent, and then generate biphenyl acetic ammonia butantriol salt. Thereaction formula is below:

In the method above, it is available to add biphenyl phenylacetic acidwith ammonia butantriol at the same time in organic solvent, or addbiphenyl phenylacetic acid first. It is also available to add ammoniabutantriol first in organic solvent, and then react with biphenylphenylacetic acid to generate biphenyl acetic ammonia butantriol salt.In order to increase recovery rate, the ratio of biphenyl phenylaceticacid and ammonia butantriol should be 1:1. As for organic solvent, thisorganic solvent should have the ability to dissolve phenyl acetic acid,such as alcohol and benzene, and especially chose methanol, anhydrousethanol, acetone, and n-butanol, benzyl and benzene. In the methodabove, there are not any special requirement for temperature and time.The normal reaction temperature is 0-80° C., but it prefers to set thetemperature at 50-70° C. The reaction time is 0.5-2 hours.

The method also includes the procedure of the purification of biphenylacetic ammonia butantriol salt. There are variety purification methodsthat we can use today, and here are two classic purification method: thefirst one is to evaporate solvent, and then generate the white powderafter the reaction. Further, the procedure is to dry the white powder invacuum, and then generate biphenyl acetic ammonia butantriol salt. Theother method is to cool down the solvent below 0° C., and then generatecrystallization. Consequently, we dry the crystallization in vacuum toproduce biphenyl acetic ammonia butantriol salt. The second method isthe best way to use, because it can get the better compound's purity andcrystal analgesic.

According to the above manufacturing method, the yield of biphenylacetic ammonia butantriol salt is 85.6-96%, and the purity is99.2-99.8%. Further, the melting point is 162-163° C.

Another object of the present invention is to provide the pharmaceuticsapplication of biphenyl acetic ammonia butantriol salt. As for theresearch, this biphenyl acetic ammonia butantriol salt hasanti-inflammatory, analgesic, and antipyretic. Accordingly, the subjectmatter of the present invention is the active compound of biphenylacetic ammonia butantriol salt.

If the purpose require, we can add one or more kinds of acceptingcarrier in this medicine, including diluting agent, excipient, extender,adhesive, moistening agent, disintegrating agent, absorbefacient,surfactant, adsorption agent, and lubricant. This medicine also can addscenting agent, sweetening agent and so on.

This medicine can be manufactured as injection (injective liquid,injective cool powder, and injective aseptic needle), tablet, powder,pill, capsule, oral liquid, cream, and so on. These above kinds ofmedicines all can be manufactured in terms of the normal order inpharmaceutical field. The amount of taking this biphenyl acetic ammoniabutantriol salt medicine is 0.8-3.6 mg/kg/days.

Still further objects and advantages will become apparent from aconsideration of the ensuing description and drawings.

These and other objectives, features, and advantages of the presentinvention will become apparent from the following detailed description,the accompanying drawings, and the appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is the HNMR spectroscopy analysis of biphenyl acetic ammoniabutantriol salt according to a preferred embodiment of the presentinvention.

FIG. 2 is the IR spectroscopy analysis of biphenyl acetic ammoniabutantriol salt according to the above preferred embodiment of thepresent invention.

FIG. 3 is the Mass spectroscopy analysis of biphenyl acetic ammoniabutantriol salt according to the above preferred embodiment of thepresent invention.

FIG. 4 shows Table 1 which illustrates solubility testing.

FIG. 5 shows Table 2 which illustrates the analgesia affection of miceheating plate method's for injecting the biphenyl acetic ammoniabutantriol salt liquid.

FIG. 6 shows Table 3 which illustrates the affection of expansion inKunming's mouse for biphenyl acetic ammonia butantriol salt injection indimethyl benzene.

FIG. 7 shows Table 4 which illustrates the affection of expansion inKunming's mouse for biphenyl acetic ammonia butantriol salt injectionglacial acetic acid.

FIG. 8 shows Table 5 which illustrates allergic reaction symptom.

FIG. 9 shows Table 6 which illustrates allergenicity evaluation standardof caviaporcellus.

FIG. 10 shows Table 7 which illustrates the active allergic testing ofanimals' change condition in caviaporcellus for injects the biphenylacetic ammonia butantriol salt injection.

FIG. 11 shows Table 8 which illustrates allergic reaction condition ofthe whole body active allergic testing in injecting the biphenyl aceticammonia butantriol salt injection.

FIG. 12 shows Table 9 which illustrates the number of 2% red bloodcell's suspending liquid.

FIG. 13 shows Table 10 which illustrates the result of hemolytic testing(application of visual) of the biphenylacetic ammonia butantriol saltinjection (high amount) (ml).

FIG. 14 shows Table 11 which illustrates the result of hemolytic testing(application of visual) of the biphenylacetic ammonia butantriol saltinjection (low amount) (ml)

FIG. 15 shows Table 12 which illustrates the exciting reaction ofrabbits' ear for the biphenyl acetic ammonia butantriol salt injection(Application of visual after 48 hours regent time).

FIG. 16 shows Table 13 which illustrates the exciting reaction ofrabbits' ear for the biphenylacetic ammonia butantriol salt injection(Application of visual after 48 hours' last reagent time).

FIG. 17 shows Table 14 which illustrates the exciting reaction ofrabbits' muscle (Application of visual) for the biphenyl acetic ammoniabutantriol salt injection (high amount).

FIG. 18 shows Table 15 which illustrates the exciting reaction ofrabbits' muscle (Application of visual) for the biphenyl acetic ammoniabutantriol salt injection (low amount).

FIG. 19 shows Table 16 which illustrates the exciting reaction ofrabbits' muscle (sickness investigation) for the biphenyl acetic ammoniabutantriol salt injection (high amount).

FIG. 20 shows Table 17 which illustrates the exciting reaction ofrabbits' muscle (sickness investigation) for the biphenyl acetic ammoniabutantriol salt injection (low amount).

FIG. 21 shows Table 18 which illustrates the test of stomach channel.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The embodiment of the present invention as shown and described below isexemplary only and not intended to be limiting. Therefore, thisinvention includes all modifications or any combination encompassedwithin the spirit and scope of the followings.

The following description illustrates a preparation of biphenyl aceticammonia butantriol salt according to a first preferred embodiment of thepresent invention.

21.2 g (0.1 ml) of Biphenyl acetate and 210 ml of methanol are addedinto a dry and clean 250 ml three opening round bottom crucible bottle.Stir and heat the mixture to 60° C. until the mixture is totallydissolved. Add ammonia butantriol into the mixture solution until thesolution is in a clarified transparent manner. Stir the solution for onehour and make TLC fully reacting with the solution. (eluent:DMF— aceticester 2:1). Set the end point when the raw material biphenyl acetate istotally reacted with another reactants. Cool down to −5° C. for 5 hoursand get huge amount of white crystal. Remove chloride and put thesolution into vacuum dry machine at 80° C. Finally, we can get thebiphenyl acetic ammonia butantriol salt in a white crystal manner. Themelting point thereof is between 162 and 163° C. The yield is 91.6%. Thetheoretical value of element analysis is C, 64.9%, H, 6.9%, N, 4.2%, O,24%. The actual value of element analysis is C, 64.1%, H, 7.3%, N, 3.6%,O, 25%).

FIG. 1 shows the ¹HNMR spectroscopy analysis of biphenyl acetic ammoniabutantriol salt.

There are eight hydrogen peaks signal in the ¹HNMR spectroscopy of thebiphenyl acetate in the solvent of DMS0-d6. According to the chemicalshift in HNMR spectroscopy, all peaks' chemical shift (δ) below 4 ppmare H signal of saturated carbon, and the total sum of peaks are 14.Therefore, if δ value is smaller than 6 ppm, there are all H signal ofaromatic ring, and the total number of these peaks is 9. At 6=3.35 ppm,the 2H single hydrogen peak cab be determined as an isolated CH₂ group.In the structure of biphenyl acetate, the carbon No. 13 which connectswith benzene and COO— in CH2 does not accept the coupling of otherhydrogen's core. At δ=3.43 ppm, the 3H single peak cab be determined asthree equal value isolated CH₂ group. In the structure of biphenylacetate, carbon NOs. 18, 19, 20 all connect with the carbon No. 17 ofthe CH₂, and these three CH₂ are equal values. Because of these threeCH₂ does not accept the coupling and have the equal value, it revealssingle peak signal. The wide peak at δ=3.43 ppm can be determined asthree OH group and a protonation's NH₃+, because these active H probablycombine with the water which is in the solvent, it reveals a wider peak.The integral value cannot explain the real number of these active H,because there is water in this solvent. There are four splitting signalgroups in the low magnetic field, and according to these peaks' area itonly can be known that there are 3, 2, 2, 2, H from high to low magneticfield. Further, there are only three left groups which can be clarifytheir peak shape, so they can be clarified as two duplicate and onetriple peaks. In addition, it can be explained that there are H on thebenzene due to their chemical shift, so two duplicate peaks near 8.0 Hzwhich named ad peak No. 7 and No. 8 can indicate as two positions. Oneposition is on a side of benzene which has H, and the other is a side ofbenzene which does not have H but have other substituted group. The twoduplicate peaks can be explained as four H in the structure of biphenylacetate which are H2/H6, H3/H5, and H8/H12. Furthermore, the triplepeaks near by 7.6 Hz can be explained as positions that those benzeneshave H on their both side, and there are H9, H10, and H11 in thismolecular structure.

IR spectroscopy analysis of biphenyl acetic ammonia butantriol salt isshown in FIG. 2.

(1) It can be determined the structure of this molecule has amine saltsof carboxylic acids, if there is a wide and strong COO— stretchingvibration peak at 1560.13 cm⁻¹, and wide, strong, and scattered NH₃+characteristic peak at the region of 2927.41-2113.60 cm⁻¹. The COO— andNH₃+ form a COO—.NH₃+ free monomer molecular structure, and COO— andNH₃+ also easily form hydrogen bonds between dimmer to polymer. Inaddition, the stretching vibration frequency region of NH₃+ and OH ofCH₂—OH reveal at 2927.41-2113.60 cm⁻¹, and then form multiple,overlapping, scattered, and strong peaks.

(2) The stretching vibration wide strong peak of OH is at 3214.94 cm⁻¹.The other deformed vibrations about OH group (60H) are at 1423.21 cm⁻¹and 1278.57 cm⁻¹, and further a flat shaking vibration is at about690.39 cm⁻¹ (rOH). It can be explained that the wide, strong, andstretching vibration peak at 1083.80 cm⁻¹ is C—O (νC—O), so it also candeclare that OH directly connect with CH₂.

(3) The series spectrum peaks at 2987˜2896 cm⁻¹, which are determinedthere are saturated carbon hydrogen group in this molecular structure,are both the asymmetric and symmetric stretching vibration group, suchas CH₃ and CH₂. At the same time, the deformed absorbing vibration ofCH₃ and CH₂ reveal at the region of 1467˜1367 cm⁻¹. Even though, it canbe reasoned from NMR spectrum that this compound does not containmethyl, there are some disrupted peaks near by 2965±cm⁻¹ and 1375±cm⁻¹from IR, so it cannot be clarify that this compound does not includemethyl.

(4) At 3100˜2980 cm⁻¹, there are a series of peaks that can be declaredthis molecular structure has unsaturated hydrocarbon group, such asaromatic ring, benzene ring, and alkene structure. At 1650˜1423 cm⁻¹,there are many unsaturated group that can be determined as C═C and C═N.At 1201˜977 cm⁻¹, it can be determined that has an internal bendingvibration (β) of ═CH group in benzene. At 738.6 and 690.39 cm⁻¹, it canbe determined that has two outer bending strong vibration peaks ofmonosubstituted benzene ring (γ). At the region of 2000-1666 cm⁻¹, aseries' peaks in this region are the same as the overlapping andcombined of a monosubstituted benzene ring and a p-disubstituted benzenering.

(5) There is stretching vibration of amine salt's C—N bond which revealsat 1045.2 cm⁻¹, and further, there is an outer bending vibration (γ) ofNH₃+ reveals at 605.5 cm⁻¹.

Mass spectroscopy analysis of biphenyl acetic ammonia butantriol salt isshown in FIG. 3.

The positive molecular ion (C₁₈H₂₃NO₅+.) of the biphenyl acetatemolecule are examined that its charge-mass ratio (m/z) is 332.2 andrelative abundance is 20. Therefore, the ratio value is 333.16 forcalculating with accurate mass theory, because the low measuringaccuracy of low resolution mass spectrometer generates mass relativeerror which is 5×10-4. In the theory, the calculating value ofC₁₈H₂₂NO₅+ ion mass fragment is 332.15 m/z, so it only has 0.6×10-4relative error.

The structure of the biphenyl acetic ammonia butantriol salt is I, andthe purification is 99.5%.

The solution test of biphenyl acetic ammonia butantriol salt:

According to Chinese medicine book in 2005 edition, the preparation ofbiphenyl acetic ammonia butantriol salt is made by different solvent. Inother words, they use acetone, formylamine, ethane nitrile, methanol,alcohol as the solvent to test their solubility. At first, mill thebiphenyl acetic ammonia butantriol salt into flour and weight it, andadd solvent into the biphenyl acetic ammonia butantriol salt. Second, at25±2° C., wave the mixer solution every 5 minutes for 30 seconds, andinspect the dissolve situation. Repeat three times and the result isillustrated in Table 1. Referring to Table 1, the biphenyl aceticammonia butantriol salt is soluble in formamide and methanol, slightlysoluble in water and alcohol, and is insoluble in acetone andacetonitrile.

Preferred Embodiment 2

21.2 g (0.1 ml) of Biphenylacetate, 12.1 g (0.1 mole) of ammoniabutantriol, and 150 ml of absolute alcohol are added into a dry andclean 250 ml three opening round bottom crucible bottle. Stir thesolution mixture at 50° C. 30 minutes for totally dissolving until thesolution is in a clarified transparent manner. Evaporate the mixturesolution and get white powder. Put the solution into vacuum dry machineat 80° C. Finally, we can get 32 grams of biphenyl acetic ammoniabutantriol salt. The melting point thereof is between 163 and 164° C.The yield is 96.0%. The theoretical value of element analysis is C,64.9%, H, 6.9%, N, 4.2%, O, 24%. The actual value of element analysis isC, 64.1%, H, 7.3%, N, 3.6%, O, 25%) The product is biphenyl aceticammonia butantriol salt, and the purity is 99.2%.

Preferred Embodiment 3

21.2 g (0.1 ml) of Biphenyl acetate and 210 ml of benzene are added intoa dry and clean 250 ml three opening round bottom crucible bottle. Stirand heat the mixture until the solution is perfect dissolved. Addammonia butantriol into the mixture solution until the solution is in aclarified transparent manner. Stir the solution for two hours and makeTLC fully reacting with the solution. Set the end point when the rawmaterial biphenyl acetate is totally reacted with other reactants judgeby TLC. Cool down to get huge amount of white crystal. Remove chlorideand use absolute ether washing the white crystal and then put it intovacuum dry machine at 80° C. Finally, we can get the biphenyl aceticammonia butantriol salt 28.5 grams. The melting point is between 163 and164° C. The yield is 85.6%. The theoretical value of element analysis isC, 64.9%, H, 6.9%, N, 4.2%, O, 24%. The actual value of element analysisis C, 64.1%, H, 7.3%, N, 3.6%, O, 25%) The product is biphenyl aceticammonia butantriol salt, and the purity is 99.8%.

Preferred embodiment 4: Preparation of the injection liquid: biphenylacetic ammonia butantriol salt.

Recipe: 94 grams of biphenyl acetic ammonia butantriol salt and 1 to 2grams of sodium carbonate to form 1000 syringes.

Use the method of preferred embodiment 1 to adjust the biphenyl aceticammonia butantriol salt into a container, add a predetermined amount ofwater. Stir the mixture and meanwhile add 0.1% (0.1 g/100 ml) sodiumcarbonate until the biphenyl acetic ammonia butantriol salt is prefectdissolved. Control the pH value between 8.5 and 8.8. Add water up to4000 ml. Add 2 gram of active carbon. Heat the solution for 15 minutes.Remove chloride and carbon from the solution. The liquid is filtered by0.22 um micro filter, and is contained and sealed it to differentsyringe tubes. (Each tube includes 94 grams of biphenyl acetic ammoniabutantriol salt). Finally, the solution is treated under a predeterminedpressure and germ-free at 115° C. to form the final product.

Preferred embodiment 5: Preparation of the dry powder of biphenyl aceticammonia butantriol salt to form 1000 dropper bottles.

Recipe: 94 grams of biphenyl acetic ammonia butantriol salt and 20 gramsof mannite.

Use the method of preferred embodiment 1 to adjust 94 grams of biphenylacetic ammonia butantriol salt into a container, add 20 grams of manniteand a predetermined amount of water for solvent. Add water up to 3000ml. Add 2 gram of active carbon. Heat the solution for 15 minutes.Remove chloride and carbon. The solution is filtered by 0.22 μm microfilters, and is contained and sealed it to different dropper bottles.(each bottle includes 94 grams of biphenyl acetic ammonia butantriolsalt).

Preferred embodiment 6: Preparation of the capsule of biphenyl aceticammonia butantriol salt.

Recipe: 188 grams of biphenyl acetic ammonia butantriol salt and 20grams of lactose to form 1000 capsules.

Use the method of preferred embodiment 2 to adjust 94 grams of biphenylacetic ammonia butantriol salt and go through 100 sieves. Addprescription lactose at 80° C. and go through 80 sieves, mixed, and testits component. Finally, fill it into the No. 1 capsule.

Preferred embodiment 7: The test of medicine effects of biphenyl aceticammonia butantriol salt.

1. Pain Effects in a Mouse Injection Biphenyl Acetic Ammonia ButantriolSalt

Pick the female Kunming mouse, each having the weight between 20 to 22grams. At first, test their sensitive towards pain. Put the mouse on thehot plate with the temperature of 55±0.5° C., and calculate how muchpain does the mouse has by determining how many timestheylick their backleg. Eliminate the mice seem to like to jump and the mice has no respondfor 30 seconds. Pick a total of 60 mice having sensitive to pain anddivide them into 5 groups randomly. Use the method of preferredembodiment 4 to adjust the biphenyl acetic ammonia butantriol salt. Thedosage of biphenyl acetic ammonia butantriol salt is divided into low,medium, and high dosage, correspondingly to 4, 12, 36 mg/kg, and normaldosage ⅓, 1, 3 respectively. And there is a positive contrast group. Themixture solution is injected into the vein of each of the mice accordingto its weight ratio 0.1 ml liquid/10 gram weight. On the other hand, thesaline is injected into the vein in the contrast group. Use epilation toremove the hair of the back leg of the mouse. Record the painful ofevery mouse before giving drug, and record the painful after giving drug1 hour. After the mouse licks its foot, remove the hot plateimmediately. Do the same record again at the second and third hour aftergiving drug. Do the t inspect, and judge its effects on easing pain.

As shown in Table 2, it shows that the pain results are very similar ondifferent mouse before giving drugs. After giving drugs, the biphenylacetic ammonia butantriol salt is significantly extends the time whichmouse is standing on the hot plate at first, second, and the third hourcompared to the contrast group. In other words, it increases the abilityof those mice to fight with pain.

2. The Pain Effects in a Kunming Mouse Injection Biphenyl Acetic AmmoniaButantriol Salt Towards their Swelling Ears.

(1) Material and Recipe:

(1.1) Test Animals:

Kunming mouse: SPF LEVEL, male, 15-17 grams, a total numbers of 110 miceare used.

Certification number: SCXK 2003-0002, 2005A012

(1.2) Drugs: Use the Method of Preferred Embodiment 4 to Adjust theBiphenyl Acetic Ammonia Butantriol Salt

2. Methods and Results

Total numbers of 110 Kunming mice are used, male, 15-17 g, medicalinspection for 3 days.

After testing, the mice are divided into eleven groups randomly and eachgroup contains 10 mice. The numbers and dosage of the biphenyl aceticammonia butantriol slat are 1, 2, 3, 4, 5 and 3, 6, 12, 24, 48,respectively. The numbers and dosage of flurbiprofenaxetil are 1, 2, 3,4, 5, and 3, 6, 12, 24, 48 respectively. Use the dimethybenzene to makethe mouse ear swelling, and use the mouse left ear as the contrastgroup. After 30 minutes, the mixing liquid is injected into the veinaccording to its weight ratio 0.1 ml liquid/10 gram weight. Use a punch,having the diameter of 8 mm, to punch the same hole in two differentears. Weight it by very high quality scale. Finally, the swelling ratiois: the weight of right ear plus the weight of left ear.

Referring to Table 3, it shows the swelling ratio of the biphenyl aceticammonia butantriol slat NO. 1, NO. 2, NO. 3, NO. 4, and NO. 5 and itsswelling ratio caused by dimethybenzene respectively. (p lower than 0.05and p higher than 0.01) On the same manner, the flurbiprofenaxetil NO.2, NO. 3, NO. 4, and NO. 5 is significantly reduces the swelling ratiotowards Kunming mouse caused by dimethybenzene.

There is no significantly difference between biphenyl acetic ammoniabutantriol and flurbiprofenaxetil. To sum up, the effect of the biphenylacetic ammonia butantriol towards the swelling ear caused bydimethybenzene is almost the same as the flurbiprofenaxetil towards theswelling ear caused by dimethybenzene.

3. The Injection of Biphenyl Acetic Ammonia Butantriol Towards the NIHMice Twisted Caused by the Acetic Acid.

1. Methods and Results

(1.1) Test Animals

NIH mice, SPF level, half of the mice are male and half of the mice arefemale, 15-17 g, a total numbers of 110 mice are used.

Certification number: SCXK 2003-0002, 2006A017

(1.2) Medicine

Use the method of preferred embodiment 4 to adjust the biphenyl aceticammonia butantriol salt.

Flurbiprofenaxetil Injection Liquid

The experiment method and result

After testing, the 110 NIH mice are divided into eleven groups randomly,which means each group has 10 mice, and each of the mice has a weightbetween 15 and 17 grams. The numbers and dosage of the biphenyl aceticammonia butantriol slat are 1, 2, 3, 4, 5 and 3, 6, 12, 24, 48 mg/kgrespectively. The numbers and dosage of flurbiprofenaxetil are 1, 2, 3,4, 5, and 3, 6, 12, 24, 48 respectively also. The medicine and weightratio is 0.1 ml medicine/10 g weight. Meanwhile, we give the samequantity of saline solution to the contrast group. After 30 minutes, themixture solution is injected into the vein according to its weight ratio0.1 ml liquid/10 gram weight for HAc painful. Record the twist reactionevery 15 minutes, count the twist reaction, and calculate the inhibitionrate.

Table 4 shows the biphenyl acetic ammonia butantriol salt solution NOs.1, 2, 3, 4, 5 and its control group can dramatically control the twistcaused by the injection glacial acetic acid. (P<0.05 or P>0.01) ED 50and 95% degree of confidence is 7.9 (4.7-13.5) mg/kg. Meanwhile, theflurbiprofenaxetil solution NOs. 1, 2, 3, 4, 5 and its control group candramatically control the twist caused by the injection glacial aceticacid. (P<0.01) ED 50 and 95% degree of confidence is 6.3 (4.9-6.3)mg/kg.

There is no significantly difference between biphenyl acetic ammoniabutantriol and flurbiprofenaxetil injection. To sum up, the effect ofthe biphenyl acetic ammonia butantriol towards the mouse twisted causedby glacial acetic acid is almost the same as the flurbiprofenaxetiltowards the mouse caused by glacial acetic acid.

Preferred embodiment 8: the safety test of injection of biphenyl aceticammonia butantriol salt.

The safety test is published by Chinese drug inspection departmentissued on August 2003. (GLP)

The Drug

(1) Use the Method of Preferred Embodiment 4 to Adjust the BiphenylAcetic Ammonia Butantriol Salt

The initial dosage to the animal: vein injection, 94 mg/injection, use0.9% NACL solution dilates to 100-200 ml, once or twice injection perday.

(2) The Positive Contrast Group: White Egg, No: 050114

(3) The Positive Contrast Group: 0.9% (0.9 g/100 ml) NACL Solution

2. Test Animal and Feed Condition:

Guinea Pig

a. level, race: the common level guinea pig.

b. Age: 10 to 12 weeks old.

c. First weight when purchasing/sex: 280-330 g, half of guinea pigs aremale and half of guinea pigs are female.

d. original from, and evidence of certification: certification number:SCXK 2006-0015, 2006B-007.

e. animal pick explanation: guinea pig is one of the priorityexperiments for allergy test, it is recommended by the drug researchguide.

f. Inspection procedure: do the investigation to the test animal beforeinspection. Review the appearance and behavior of the animal includingdefecation, food intake, and so on, and eliminate viral animal. Theentire animal doing the investigation are required to have the permitfrom the investigation person.

g. Standard method: use the drying method, and use saturated sorrow andbitter solution to mark spot on different parts of animal for differentnumber. Each group has 3 guinea pigs, and mark them respectively.

2. Rabbit

a. level, race: the common level rabbit.

b. Age: 3 months old.

c. First weight when purchasing, sex: 2.0-2.2 kg, half of the rabbitsare male and half of the rabbits are female.

d. original from, and evidence of certification: certification number:2006A001

e. animal pick explanation: rabbit is one of the priority experiments ofallergy test, it is recommended by the drug research guide.

f. Inspection procedure: the procedure is the same as guinea pig does.

g. Standard method: the method is the same as rabbit does. Feed therabbits separately, and mark them.

(3) Feed Condition:

a. Feed management: the animal is feeding by people who own thecertificate of feeding animal.

b. Feed condition, level, and certification: guinea pig and the rabbitare living in common guinea pig room and rabbit room respectively, allthe utility meets the requirement of safety animal living, and good airventilation and light, temperature is between 16-26° C., humidity is 40to 70%, The animal safety certification number is: 2005C133. The testanimal approval number is: SYXK-2003-0003 (ISSUED BY 2005).

c. Feed and drinking water: drinking water is the stuffs of feed.

Use the method of preferred embodiment 4 to adjust the biphenyl aceticammonia butantriol salt and to do the safety experiment.

No. 1 the biphenyl acetic ammonia butantriol salt injection towards theguinea pig for the whole body allergy experiment

1. Method:

Take 24 experimental guinea pig, evenly divide into 4 groups by sex andweight randomly, each group has six male guinea pig and six femaleguinea pig. It divided into the positive contrast group, the negativecontrast group, the low and high dosage group from the method ofpreferred embodiment 4 to adjust the biphenyl acetic ammonia butantriolsalt, and it is 7.5 and 22.5 mg/kg bw equaling to one times and threetimes of the clinical usage respectively.

Injection through abdominal cavity, and the dosage per animal is 0.5 ml,for every other day for three sequence injections. Sensitizer dosage:the positive contrast group: NaCl solution is given (0.9 g/ml); thepositive contrast group: 2% (2 g/100 ml) white egg is given; the lowdosage group: 7.5 mg the biphenyl acetic ammonia butantriol salt/kg bw.The high dosage group: 22.5 mg the biphenyl acetic ammonia butantriolsalt/kg bw.

The concentration of medicine: the low dosage group: 5.0 mg biphenylacetic ammonia butantriol salt/ml; the high dosage group: 15.0 mgbiphenyl acetic ammonia butantriol salt/ml, wherein the method of makingthe high dosage solution is that use 1 bottle of biphenyl acetic ammoniabutantriol salt solution (4 ml: 94 mg/bottle) from the method ofpreferred embodiment 4. Take 4 ml the biphenyl acetic ammonia butantriolsalt solution and dilute it with 0.9% NACL solution to 6.3 ml. Theconcentration of the biphenyl acetic ammonia butantriol salt solutionis: 15.0 mg the biphenyl acetic ammonia butantriol salt solution/mlsolution). The method of making the low dosage solution is that use 1bottle of biphenyl acetic ammonia butantriol salt solution (2 ml: 15mg/bottle) from the method of preferred embodiment 4. Take 2 ml thebiphenyl acetic ammonia butantriol salt solution and dilute it with 0.9%NaCl (0.9 g/100 ml) solution to 6.0 ml. The concentration of thebiphenyl acetic ammonia butantriol salt solution is: 5.0 mg the biphenylacetic ammonia butantriol salt solution/ml solution).

After 12 days of the cavity injection, the medicine is given by veininjection. medicine volume: 1.0 ml/per animal. Dosage: two times ofallergy dosage. Inspection the animal responds.

2. Target of Investigation:

2.1 Allergy Period: Weigh the Animal Before Allergy, and Record theAnimal Responds in Detail.

2.2 Allergic Reaction: Weight the Animal Before Exciting. Inspect theAnimal Responds for 30 Minutes and Record in Table 5 in Detail.

3. Result:

We can judge the level of allergy and the reaction rate of allergy. Ifthere is any animal having allergy after rejection, we can take twohealthy guinea pigs to do the vein injection for medicine 2 ml. Do theInspection of that are there any allergy situation caused by the feedingmaterial.

4. Result:

4.1 Investigation in Normal Condition

In the period of investigation, the activity, the breath, and the stoolexamination of the guinea pigs have not showed significantly unusual.The weight change is shown in Table 7.

4.2 Active Allergic Reactions:

The allergic action and the active allergic towards the guinea pigs areshown in Table 8. There is no allergy symptom occurred in the negativecontrol group. There is allergy symptom occurred in the positivecontrol. The symptoms for six guinea pigs are hyperphea, polypena, stepunstable, jumping, and death. There is no allergic symptom in low dosagereagent group and the high dosage reagent group.

The results show that there is no allergy symptom in the negativecontrol group, but the entire guinea pigs of the positive control groupwent death. There is no allergic symptom in low dosage reagent group andthe high dosage reagent group.

The results show that we use the method of preferred embodiment 4 toadjust the biphenyl acetic ammonia butantriol salt and do the whole bodyallergy test towards the guinea pig is negative effect.

2. The Blood Absorbing Test Towards the Biphenyl Acetic AmmoniaButantriol Salt

The blood absorbing test outside the body: put the biphenyl aceticammonia butantriol salt making from the method of preferred embodiment 4solution 0.63/mg/ml, 1.88 mg/ml into 2% red blood cell's suspendingliquid, respectively. There is no blood absorbing reaction within 3hours in each tube. The results show that we use the method of preferredembodiment 4 to adjust the biphenyl acetic ammonia butantriol salt anddo the absorbing test outside the body towards the guinea pig isnegative effect. The detail experimental methods and results are shownin the following:

The method of making the medicine:

(1) high dosage group: the high dosage group: use 1 bottle of biphenylacetic ammonia butantriol salt solution (4 ml: 94 mg/bottle) from themethod of preferred embodiment 4. Take 0.5 ml the biphenyl aceticammonia butantriol salt solution and dilute it with 0.9% NaCl solution(0.9 g/100 ml) to 6.25 ml. The concentration of the biphenyl aceticammonia butantriol salt solution is: 1.88 mg/the biphenyl acetic ammoniabutantriol salt solution/ml solution).

(2) low dosage group: use 1 bottle of biphenyl acetic ammonia butantriolsalt solution (4 ml: 94 mg/bottle) from the method of preferredembodiment 4. Take 2 ml the biphenyl acetic ammonia butantriol saltsolution and dilute it with 0.9% NACl (0.9 g/100 ml) solution to 6.0 ml.The concentration of the biphenyl acetic ammonia butantriol saltsolution is: 0.63 mg the biphenyl acetic ammonia butantriol saltsolution/ml solution).

2. The Injection Method:

(1) The Preparation of 2% Red Blood Cell's Suspending Liquid

Take few milliliters of rabbit blood into a triangular flask and shakeit for 10 minutes. Remove the fiber egg white, and get non-fiber blood.Separate the blood solution into few centrifuge tubes. Add ten times of0.9% NaCl solution, well mixing, and centrifuge (1500 turns/minutes, 15minutes) to remove the supernatant. Use 0.9% NaCl solution to wash theprecipitation red blood two or three times until the supernatant is notred anymore. Use the red blood to mix the 0.9% NaCl solution to make 2%suspending liquid for test use.

Take the same size of opening of 7 clean test tubes and number them. Addthe 0.9% NaCl solution, the injection water, medicine, and the 2%suspending liquid followed by Table 9. After well mixing, put them inthe temperature chamber at 37° C. Inspect them every 15 minutes. After 1hour, inspect them every one hour for 3 hours.

(2) Result Observation

If testing solution presents clearly red, no cellular residual under thetube, and few cellular residual, it can determines that haemolysishappens. If red cells all sink, and supernatant fluid presents no color,it can present that haemolysis doesn't happen. If there are red brownprecipitates in the solution, and non-dispersed after shaking, itrepresents that the red cellular condensed matter happens. If redcellular coagulation happens, and further, it must be decide that it istrue or faked coagulation. If condensed matters are uniform distributionafter shaking the tube, it can be decided as faked coagulation. Orputting aggregate on this wave plate and put 2 drops 0.9% NaCl injectionon the fringe of cap wave plate, and then observe in the scope, if redcellular condensed matter can be flushed scattering, it is fakedcoagulation. If condensed matter cannot be flushed scattering, it istrue coagulation.

3. Result Decision

While negative control tube has no haemolysis and coagulation, positivecontrol tube has haemolysis, and the subject tube does not hashaemolysis and coagulation in three hours, this sample (tested animal)can be used for injecting. If the subject tube has haemolysis andcoagulation in three hours, this sample cannot be used for injecting.

4. The Results

After adding low concentration 0.63 mg/ml and high concentration 1.88mg/ml the biphenyl acetic ammonia butantriol salt into different tubes,there is no blood absorbing reaction within 3 hours. In other words, theoutside body blood test is negative effect as shown in Table 10 andTable 11.

The blood test of the biphenyl acetic ammonia butantriol salt injectionsolution

The test of the blood of rabbit's ear

Pick 8 healthy rabbits; use the contrast method towards each right andleft ear. Vein injection to the rabbit's left ear. The concentration ofthe medicine is 5 ml/kg weight. Use the method of preferred embodiment 4to adjust the biphenyl acetic ammonia butantriol salt solution. The lowdosage and high dosage of the biphenyl acetic ammonia butantriol saltsolution is 3.15 mg/kg bw, and 9.4 mg/kg bw, respectively. Meanwhile theconcentration of them is 0.63 mg/ml, and 1.88 mg/ml respectively whichis 0.7-1.4 and 2-4 times of clinical vein injection. Vein injection tothe rabbit's right ear for 0.9 g/100 ml NaCl is formed as the controlgroup. Once a day, and repeats it for consecutive three days. Afterinjection high and low dosage of solution, each rabbits is given 0.9%NACL liquid. Pick each two given high dosage and low dosage rabbits, anddissect it after 48 hours. Giving another two given high dosage and lowdosage rabbits for two weeks recovery period, and then dissect it too.Finally, a sharp outline of a blood vessel, the thickness of rabbit'sear is even. There is no significant difference after using. There is noany poison on the animal's blood according to pathology. The method ofpreferred embodiment 4 to adjust the biphenyl acetic ammonia butantriolsalt solution is conforming to the regulation standard.

The method of the experiment and the result is showing on the following:

1. The Preparation of Test:

(1) High dosage group: use 2 bottles of biphenyl acetic ammoniabutantriol salt solution (4 ml: 94 mg/bottle) from the method ofpreferred embodiment 4. Take 8 ml the biphenyl acetic ammonia butantriolsalt solution and dilute it with 0.9% NaCl solution (0.9 g/100 ml) to100 ml. The concentration of the biphenyl acetic ammonia butantriol saltsolution is: 1.88 mg/the biphenyl acetic ammonia butantriol saltsolution/ml solution).

(2) Low dosage group: take 30 ml of the above solution havingconcentration of 1.88 mg/ml. Take 30 ml of the biphenyl acetic ammoniabutantriol salt solution and dilute it with 0.9% NaCl (0.9 g/100 ml)solution to 90 ml. The concentration of the biphenyl acetic ammoniabutantriol salt solution is: 0.63 mg the biphenyl acetic ammoniabutantriol salt solution/ml solution).

2. Weight the Animal: Weight the Animal Before Injection, and afterInjection for 48 Hours and 14 Days.

3. Regular Inspection and Animal Pick

Observe and record the response of the animal every day before injectionevery day. After the final injection for 48 hours, the highconcentration and the low concentration rabbits are being killed bybleeding. Record the blood reaction. Cut the left ear first, and thencut the right ear from the boundary part of the ear, and then mark it.Cut a portion of the ears sample into 10% methylene liquid. The sampleis 8 cm in length, and 1 cm in width, wherein a first cut is formed at0.5 cm from the first needle point, a second cut is formed at 2 cm fromthe third needle point, and the thread side is the second cut. On theother hand, after 14 days of giving medicine, the high and low dosageanimals do the following inspection. Use the first needle point as theboundary, cut one section from the first cut. And use the third needlepoint as boundary, cut two sections from the second cut. Use paraffin toform the sampling slice. The thickness of sampling slice is between 4and 5 μm. The sampling slice is treated by H-E dying for inspection.

4. Judge the Results

Do the comprehensive judge by visual examination and inspection.

5. The Results

5.1 Visual Examination:

Observe and record the response of the animals every day beforeinjection. Between the drugs given periods, we can see the skin of therabbit's ear for the high concentration and low concentration is red.The size is between 0.1×0.2 and 0.2×1.0 cm. After 48 hours of finalinjection, the investigation of four rabbits of high concentration andlow concentration: a sharp outline of a blood vessel, the thickness ofrabbit's ear is even. There is no significant difference after using asshown in Table 12 and Table 13. After 14 days of final injection, theinvestigation of four rabbits of high concentration and lowconcentration: a sharp outline of a blood vessel, the thickness ofrabbit's ear is even. There is no significant difference after using.

5.2 The Investigation:

After 48 hours of giving high and low dosages of the biphenyl aceticammonia butantriol salt solution, we dissect the rabbits. On the samemethod, after two weeks of giving high and low dosages of the biphenylacetic ammonia butantriol salt solution, we dissect the rabbits. Thereis no degenerate and necrosis on the structure of the blood.

The muscle test of the biphenyl acetic ammonia butantriol salt solution

Pick 8 healthy rabbits; use the contrast method towards rabbit's muscle.Vein injection to the rabbit's left muscle once a day for 3 days. Theconcentration of the medicine is 1 ml/kg weight. Use the method ofpreferred embodiment 4 to adjust the biphenyl acetic ammonia butantriolsalt solution. The concentration of low dosage and high dosage of thebiphenyl acetic ammonia butantriol salt solution is 0.63 mg/ml, and 1.88mg/ml respectively which is 0.7-1.4 and 2-4 times of clinical veininjection. Vein injection to the rabbit's right muscle for 0.9 g/100 mlNACL is set as the control group. After injection of high and lowdosages of solution, each rabbits is given 0.9% NaCl liquid. Pick eachtwo given high dosage and low dosage rabbits, and dissect it after 48hours. Giving another two given high dosage and low dosage rabbits twoweeks recovery period, and then dissect them too. Do the visualinspection of eight rabbits: the muscular tissue around injection isfull of flexibility and luster, no significant change. There is nodegenerate and necrosis on the structure of the deep muscle of therabbit according to pathology. The method of preferred embodiment 4 toadjust the biphenyl acetic ammonia butantriol salt solution isconforming to the regulation standard.

The method of the experiment and the result is showing on the following:

1. The Preparation of Test:

(1) High dosage group: use 1 bottle of biphenyl acetic ammoniabutantriol salt solution (4 ml: 94 mg/bottle) from the method ofpreferred embodiment 4. Take 1 ml the biphenyl acetic ammonia butantriolsalt solution and dilute it with 0.9% NaCl solution (0.9 g/100 ml) to12.5 ml. The concentration of the biphenyl acetic ammonia butantriolsalt solution is: 1.88 mg/the biphenyl acetic ammonia butantriol saltsolution/ml solution).

(2) Low dosage group: take 2 ml of the above solution havingconcentration 1.88 mg/ml. Take 2 ml the biphenyl acetic ammoniabutantriol salt solution and dilute it with 0.9% NaCl (0.9 g/100 ml)solution to 6 ml. The concentration of the biphenyl acetic ammoniabutantriol salt solution is: 0.63 mg the biphenyl acetic ammoniabutantriol salt solution/ml solution).

2. Weight the Animal: Weight the Animal Before Injection, and afterInjection for 48 Hours and 14 Days.

3. The Injection Method

Pick 8 healthy white rabbits. Inject high dosage and low dosage biphenylacetic ammonia butantriol salt solutions into left muscle using asepticmethod. On the contrary, inject the same volume 0.9% NaCl into the rightmuscle as the contrast group.

4. Regular Inspection

Observe and record the injection portions around the muscle portions ofthe animals every day before injection. After the final injection for 48hours, the high concentration and the low concentration rabbits arebeing killed by bloodletting, and expose the muscle. Cut it throughlongitudinal direction. Record the injection portion by visualexamination. On the other hand, after 14 days of giving medicine, tworabbits were given by the high and low dosage medicine animal do theinspection around the injection portion.

5. Judge the Results

Do the comprehensive judge by visual examination and inspection.

6. The Results

The result is shown in the Table 14, Table 15, Table 16, Table 17, andthe diagnosis report.

After 48 hours and 14 days of giving high and low dosage of the biphenylacetic ammonia butantriol salt solution, we do the visual examinationand read the report: After using the biphenyl acetic ammonia butantriolsalt solution on the rabbit's muscle portion, there is no degenerate andnecrosis on the structure of the muscle portion.

The results show that we use the method of preferred embodiment 4 toadjust the biphenyl acetic ammonia butantriol salt (7.5/22.5 mg/kg bw)and do the whole body allergy test towards the guinea pig is negativeeffect. The results show that we use the method of preferred embodiment4 to adjust the biphenyl acetic ammonia butantriol salt 0.63/1.88 mg/ml)and do the inspection towards the rabble's ear boundary is negativeeffect. The results show that we use the method of preferred embodiment4 to adjust the biphenyl acetic ammonia butantriol salt 0.63/1.88 mg/ml)and do the inspection towards the rabbit for outside body bloodabsorption is negative effect.

Preferred embodiment 9: The poison test towards the mouse for thebiphenyl acetic ammonia butantriol salt

We cannot judge the effects of the biphenyl acetic ammonia butantriolsalt injection towards the NIH mice (half mousse are died) Therefore, weuse the maximum dosage of the biphenyl acetic ammonia butantriol saltinjection towards the NIH mice to do the research about the poison ofthe biphenyl acetic ammonia butantriol salt towards the NIH mice.

A total amount of 40 NIH mice, 19-21 gram, (half of the mice are maleand half of the mice are female), it is divided into two groups:medicine group and the contrast group. The medicine is given by themaximum dosage of the biphenyl acetic ammonia butantriol salt (94 mg/4ml) medicine volume: 0.2 ml/once/10 g weight), twice a day, twoinjections are six hours difference, i.e. the total dosage for one dayis 940 mg/kg. The contrast is given by the 0.1% NaCl liquid.Investigation and record the poison effect on the mice and the amount ofmice death. The result is that the entire mice in the medicine group arebarely to move. Some mice are very hard to breath. After injection for 2to 4 hours, the condition of the entire mice is getting better. Theentire condition of the mice in the contrast group is normal. Afterinjection for 14 days, no mouse went death in both group, and they lookgood for gaining weight. And dissect all of them; there is nosignificantly change by visual examination. The research show that themaximum enduring quantity of the NIH mice towards the biphenyl aceticammonia butantriol salt is over 940 mg/kg. (calculated for the enduringquantity for biphenyl acetic ammonia is 598 mg/ml) Meanwhile biphenylacetic acid ester for vein injection LD50 is 337 mg/ml for male, and 433mg/ml for female. The position of the biphenyl acetic ammonia butantriolis significantly decreased, and its poison is also lower than otheranti-inflammatory agent.

Preferred embodiment 10: the stable test of the biphenyl acetic ammoniabutantriol salt injection

According to the Chinese pharmacopia appendix XI X C, second edition,2005 (the rule of medicine stability testing direction), Committing theimplement sample 4 of the stability investigation item for biphenylacetic ammonia butantriol salt injection includes the influence factorstesting (includes high temperature, high humidity, and highlights),accelerating testing, long term testing and so on. The purpose is toinvestigate the regular change of this product which has the influenceof temperature, humidity, and light intensity, and offer producing,packaging, and transporting condition for this medicine. At the sametime, the effective date is established by passing this testing.

According to the Chinese pharmacopia appendix XI X C, second edition,2005 (the rule of medicine stability testing direction) and “Theimportant stability item-list of bulk drug and pharmaceuticalpreparation” which is in “The researching directive rule of chemicalmedicine and therapeutic biological products” determines the importantitem-list, which includes gender character, PH value, relativematerials, asepsis, thermal source, assay, and so on. The testing resultis below:

1. Influence Test:

After putting the biphenyl acetic ammonia butantriol salt at hightemperature 60° C., high humidity (relative humidity 92.5±5%), and lightintensity (4500±5001×) for 10 days, there is no significantly change forthe product in different standard evolution. In other words, the qualityof the product is very stable.

2. Acceleration Test

After putting the biphenyl acetic ammonia butantriol salt at hightemperature 40±2° C., high humidity (relative humidity 75±5%), and for 3months, there is no significantly change for the product in every monthexamination. In other words, the quality of the product is very stable.

3. Long Term Test

After putting the biphenyl acetic ammonia butantriol salt at hightemperature 25° C., high humidity (relative humidity 25%) for 3 months,there is no significantly change for the product in every monthexamination. In other words, the quality of the product is very stable.

Preferred embodiment 11: the test of stomach channel towards thebiphenyl acetic ammonia butantriol salt oral administration

NIH mice are divided into two groups randomly. After fasting for 24hours, the NIH mice are fed by 333 mg/kg of the biphenyl acetic ammoniabutantriol salt, and 212 mg/kg of the biphenyl acetic. Dissect stomachafter 48 hours. Use 10% methylene oxide. Cut the stomach along the curveof stomach, and calculate the percentage of an ulcer canker. The resultis shown in Table 18.

Industry Application

The biphenyl acetic ammonia butantriol salt of the present invention candissolve in water, so this benefit can solve the problem of incapable ofdissolving the biphenyl phenylacetic acid in water. At the same time ofsolving the solubility problem, the present invention proves theanalgesic and anti-inflammatory equivalence of biphenyl acetic ammoniabutantriol salt and biphenyl phenylacetic ester. In addition, we canknow the result is that biphenyl acetic ammonia butantriol salt has thelower acute toxicity than biphenyl phenylacetic ester (see Example 9),and lower oral irritation (see Example 11). On this basis, we go a stepfurther to do the allergic reactive experiment which follows the GLPtesting rule, and finally, we don't find the allergic reaction andshocking phenomena which cause from allergy. At the same time, accordingto a normal injection requirement, we did the haemolysis and irritatingtesting of biphenyl acetic ammonia butantriol salt, and then we foundthat there is no irritation of haemolysis, muscle, and blood vessel, soit can use for injection.

All of the above experiment proceeds in the national new medicine safetyevaluation pharmacy research important laboratory, so the data have thehigh reliability. This medicine can be used for injection, and avoid touse big molecular coemulsifier of producing emulsion. If we want to usebiphenyl phenylacetic ester for emulsion, stability is the big problemfor pharmaceutics workers, such as acidification. The present inventiondoes a three months stability testing of biphenyl acetic ammoniabutantriol salt injection, and we find that biphenyl acetic ammoniabutantriol salt injection has high stability. The procedure of producingemulsion is re-staining, but the procedure of producing biphenyl aceticammonia butantriol salt and coemulsifier is easy to proceed.

The manufacturing technique of biphenyl acetic ammonia butantriol saltis very easy to proceed. The raw material is very easy to find, and thereaction is stability, high yield, and the product purity is high. So,we can directly to get the medical biphenyl acetic ammonia butantriolsalt. In addition, there is no serious environmental pollutant in thisproducing procedure. According to the pharmaceutical testing, it can beproved that biphenyl acetic ammonia butantriol salt injection has theinfluence to ease analgesic, anti-inflammatory, and no allergy for mice.

For the external testing, there is no red blood cell haemolysisinfluence on Japan big ears' rabbit. Because of no obviously irritantinfluence on giving many times and one time of medicine to rabbits' veinvessel and muscle, it explain that biphenyl acetic ammonia butantriolsalt can apply to muscle and vein injection. After giving one timemedicine to rabbits' vein, the result of acute toxicity testing revealsthat the tolerance value of NIH mice in injecting biphenyl aceticammonia butantriol salt is 940 mg/kg. This result reveals the LD50 valueof biphenyl acetic ammonia butantriol salt is higher than 940 mg/kg(biphenyl phenylacetic acid is 589 mg/kg), but the LD50 value ofbiphenyl phenylacetic ester is 337 mg/kg for male and 433 mg/kg forfemale. The acute toxicity of biphenyl acetic ammonia butantriol saltdramatically decreases, and its acute toxicity is lower than the samekinds of anti-inflammatory and analgesics medicines. Biphenyl aceticammonia butantriol salt has the lower irritant of stomach, intestine,mouth, and abdomen than biphenyl phenylacetic acid, and further easy totake this medicine.

Accordingly, the present invention provides A composition for providinganti-inflammatory, analgesic, and antipyretic effect, wherein thecomposition is a biphenyl acetic ammonia butantriol salt having achemical structure of:

wherein the biphenyl acetic ammonia butantriol salt has a molecular massof 333.14. The biphenyl acetic ammonia butantriol salt is soluble inmethanol and methanamide, slightly soluble in water and ethanol, andinsoluble in propanol and acetonitrile. It is worth mentioning that thebiphenyl acetic ammonia butantriol salt has low toxicity to and lowstimulative effect on a living object. In addition, the biphenyl aceticammonia butantriol salt is stable in structure and is non-allergic tothe living object, thereby the biphenyl acetic ammonia butantriol saltis capable of being prepared into an injection form for administrationby injection without causing allergy to the living object.

The biphenyl acetic ammonia butantriol salt is prepared by a method ofpreparation comprising the steps of: mixing biphenyl phenylacetic acidand ammonia butantriol in organic solvent to produce biphenyl aceticammonia butantriol salt, wherein a ratio of the biphenyl phenylaceticacid and the ammonia butantriol is 1:1. It is worth mentioning that thebiphenyl acetic ammonia butantriol salt produced by the method ofpreparation has a purity of more than 99.5% by weight.

The reaction temperature is set at 0-80° C., and is preferably at 50-70°C. The reaction time is set as 0.5-2 hours.

Preferably, the biphenyl phenylacetic acid is firstly dissolved in theorganic solvent, and then the ammonia butantriol is added into theorganic solvent for chemical reaction to generate the biphenyl aceticammonia butantriol salt. The ratio of the biphenyl phenylacetic acid andthe ammonia butantriol is 1:1.

Preferably, the organic solvent is selected from the group consisting ofalcohol group and benzene group. Preferably, the alcohol group isselected from the followings: methanol, anhydrous ethanol, acetone, andn-butanol. Preferably, the benzene group is selected from the groupconsisting of benzyl and benzene.

Preferably, the method of preparation further comprises the step ofpurifying the biphenyl acetic ammonia butantriol salt such that a purityof the biphenyl acetic ammonia butantriol salt is greater than 99.5%.

In particular, the method further comprises the steps of: evaporating amixture of the biphenyl phenylacetic acid and the ammonia butantriol toobtain white powder after purification, and drying the white powder in avacuum condition to obtain the biphenyl acetic ammonia butantriol salt.Alternately, the method further comprises the steps of: crystallizing amixture of the biphenyl phenylacetic acid and the ammonia butantriol ata temperature below 0° C. to form a crystal of the mixture, and dryingthe crystal in a vacuum condition to obtain the biphenyl acetic ammoniabutantriol salt.

It is worth mentioning that the method may further include the steps of:preparing the biphenyl acetic ammonia butantriol salt into an injectionform adapted for injection to a living object, wherein the biphenylacetic ammonia butantriol salt in the injection form is stable instructure and has insignificant local and vascular irritating effectsuch that the injection form is capable of providing anti-inflammatory,analgesic, and antipyretic effect by injecting into a living object,thereby the biphenyl acetic ammonia butantriol salt having a unique andstable chemical structure is capable of being prepared into apredetermined dosage in a controllable and precise manner. The injectionform includes injection liquid, injection cool powder and injectionaseptic needle.

On the other hand, the method may further include the steps of:preparing the biphenyl acetic ammonia butantriol salt into a capsuleform adapted for oral administration, wherein the biphenyl aceticammonia butantriol salt in the capsule form is stable in structure andhas insignificant local and vascular irritating effect such that thebiphenyl acetic ammonia butantriol salt is capable of providinganti-inflammatory, analgesic, and antipyretic effect to a living object,thereby the biphenyl acetic ammonia butantriol salt having a unique andstable chemical structure is capable of being prepared into apredetermined dosage in a controllable and precise manner.

It will thus be seen that the objects of the present invention have beenfully and effectively accomplished. It embodiments have been shown anddescribed for the purposes of illustrating the functional and structuralprinciples of the present invention and is subject to change withoutdeparture from such principles. Therefore, this invention includes allmodifications encompassed within the spirit and scope of the followingclaims.

1. A composition for providing anti-inflammatory, analgesic, andantipyretic effect, wherein the composition is a biphenyl acetic ammoniabutantriol salt having a chemical structure of:

wherein said biphenyl acetic ammonia butantriol salt has a molecularmass of 333.14.
 2. The composition according to claim 1, wherein saidbiphenyl acetic ammonia butantriol salt is soluble in methanol andmethanamide, slightly soluble in water and ethanol, and insoluble inpropanol and acetonitrile, wherein said biphenyl acetic ammoniabutantriol salt has low toxicity to and low stimulative effect on aliving object.
 3. The composition according to claim 2, wherein saidbiphenyl acetic ammonia butantriol salt is stable in structure and isnon-allergic to the living object, thereby said biphenyl acetic ammoniabutantriol salt is capable of being prepared into an injection form foradministration by injection without causing allergy to the livingobject.
 4. The composition according to claim 3, is prepared by a methodof preparation comprising the steps of: mixing biphenyl phenylaceticacid and ammonia butantriol in organic solvent to produce biphenylacetic ammonia butantriol salt, wherein a ratio of the biphenylphenylacetic acid and the ammonia butantriol is 1:1.
 5. The compositionaccording to claim 4, wherein said biphenyl acetic ammonia butantriolsalt produced by said method of preparation has a purity of more than99.5% by weight.
 6. A method of preparation of biphenyl acetic ammoniabutantriol salt, comprising the steps of: mixing biphenyl phenylaceticacid with ammonia butantriol in organic solvent to generate biphenylacetic ammonia butantriol salt, wherein biphenyl acetic ammoniabutantriol salt having a chemical structure of:


7. The method according to claim 6, wherein said biphenyl phenylaceticacid is firstly dissolved in said organic solvent, and then said ammoniabutantriol is added into said organic solvent for chemical reaction togenerate said biphenyl acetic ammonia butantriol salt.
 8. The methodaccording to claim 6, wherein a ratio of said biphenyl phenylacetic acidand said ammonia butantriol is 1:1.
 9. The method according to claim 7,wherein a ratio of said biphenyl phenylacetic acid and said ammoniabutantriol is 1:1.
 10. The method according to claim 8, wherein saidorganic solvent is selected from the group consisting of alcohol groupand benzene group.
 11. The method according to claim 10, wherein saidalcohol group of said organic solvent is selected from the groupconsisting of methanol, anhydrous ethanol, acetone, and n-butanol,wherein said benzene group of said organic solvent is selected from thegroup consisting of benzyl and benzene.
 12. The method according toclaim 10, wherein a reaction temperature is set at 0-80° C. and areaction time is set as 0.5-2 hours.
 13. The method according to claim12, wherein a reaction temperature is set at 50-70° C.
 14. The methodaccording to claim 8, further comprising a step of purifying saidbiphenyl acetic ammonia butantriol salt such that a purity of saidbiphenyl acetic ammonia butantriol salt is greater than 99.5%.
 15. Themethod according to claim 12, further comprising a step of purifyingsaid biphenyl acetic ammonia butantriol salt such that a purity of saidbiphenyl acetic ammonia butantriol salt is greater than 99.5%.
 16. Themethod according to claim 13, further comprising the steps ofevaporating a mixture of said biphenyl phenylacetic acid and saidammonia butantriol to obtain white powder after purification, and dryingsaid white powder in a vacuum condition to obtain said biphenyl aceticammonia butantriol salt.
 17. The method according to claim 13, furthercomprising the steps of crystallizing a mixture of said biphenylphenylacetic acid and said ammonia butantriol at a temperature below 0°C. to form a crystal of said mixture, and drying said crystal in avacuum condition to obtain said biphenyl acetic ammonia butantriol salt.18. The method according to claim 15, further comprising the steps of:preparing said biphenyl acetic ammonia butantriol salt into an injectionform adapted for injection to a living object, wherein said biphenylacetic ammonia butantriol salt in said injection form is stable instructure and has insignificant local and vascular irritating effectsuch that said injection form is capable of providing anti-inflammatory,analgesic, and antipyretic effect by injecting into a living object,thereby said biphenyl acetic ammonia butantriol salt having a unique andstable chemical structure is capable of being prepared into apredetermined dosage in a controllable and precise manner.
 19. Themethod according to claim 18, wherein said injection form is selectedfrom the group consisting of injection liquid, injection cool powder andinjection aseptic needle.
 20. The method according to claim 15, furthercomprising the steps of: preparing said biphenyl acetic ammoniabutantriol salt into a capsule form adapted for oral administration,wherein said biphenyl acetic ammonia butantriol salt in said capsuleform is stable in structure and has insignificant local and vascularirritating effect such that said biphenyl acetic ammonia butantriol saltis capable of providing anti-inflammatory, analgesic, and antipyreticeffect to a living object, thereby said biphenyl acetic ammoniabutantriol salt having a unique and stable chemical structure is capableof being prepared into a predetermined dosage in a controllable andprecise manner.